1. Field of Invention
This invention relates generally to techniques for healing lesions in cartilage, and more particularly to a technique in which lesions in articular cartilage are healed by using in vitro cultured chondrocytes as autografts.
2. Status of Prior Art
The concern of the present invention is with cartilage, an important constituent of which is collagen. Hence by way of introduction, we shall first briefly consider the nature of collagen, a substance which accounts for about thirty percent of the total human body protein. Collagen, which has a characteristic amino acid composition, forms the fibrillar component of soft connective tissues such as skin, ligament and tendon, and is the major component of the organic matrix of calcified hard tissues such as bone and dentine. Quite apart from its structural significance, collagen plays an important role in development and in wound healing.
There exist at least four genetically distinct types of collagen. The most familiar, type I, consists of three polypeptide chains. Two chains are identical and are called .alpha.1(I); the third being called .alpha.2. Type I collagen forms the major portion of the collagen of both soft (skin, tendon and hard (bone and dentine) connective tissue. Type II collagen is the major collagen of cartilage and is composed of three .alpha.1(II) chains. Type III collagen is composed of three .alpha.1(III) chains and is found in blood vessels, wounds, and certain tumors. Reticulin fibers appear to be identified with type III collagen. Basement membrane collagens are classified as type IV.
The intercellular substances of connective tissue are classified as either amorphous or fibrous. The former exist as firm or soft gels. These are mucopolysaccharides containing bound water which permits diffusion to take place through them. Fibrous intercellular substances are commonly immersed in the amorphous type and assume various forms, such as the white fibers of collagen, yellow fibers constituting an elastin, and reticular fibers in the form of lacy networks that give intimate internal support to cells. Connective tissue cells that generate intercellular substances thereafter lie within the substances they have formed.
Cartilage, which is a specialized kind of connective tissue, serves as the model for most bones during development, and it persists in adults in certain limited regions in three forms: hyaline or glassy cartilage; elastic cartilage and fibrocartilage.
Hyaline cartilage is found in joints, at the ventral ends of ribs and in other regions of the body. The cartilage cells or chondrocytes take up about one third of the volume of the cartilage, the rest being occupied by the matrix which separates the cells and contains collagen fibers. Since cartilage has no blood vessels, exchanges with blood occur over longer distances than in other forms of connective tissue.
Elastic cartilage, which is found in the external ear, in the epiglottis and at other sites, is more opaque and flexible than hyaline cartilage and is rich in elastic fibers. Fibrocartilage, which occurs in certain tendons near their attachment to bones, is constituted by dense bundles of collagen fibers, the cells being disposed in columns between these fibers.
The intercellular substance of cartilage is chiefly composed of a mucopolysaccharide gel and collagen fibers. The cells which create this intercellular substance are called chondoblasts, and after making this substance they lie within it as chondrocytes.
Appended to this specification is a listing of cited prior art publications which identify the authors, the published papers and their publishers, and also give the relevant page numbers. In the discussion to follow, these papers will be referred to only by their authors and year of publication.
Articular cartilage is classified as a connective tissue along with bone and ligaments (Bloom and Fawcett, 1975). Articular cartilage combines two components; namely, a cellular component comprising the chondrocytes, and an extracellular matrix which consists of collagen and proteoglycan. Articular cartilage is found at the articulating ends of most diarthodial synovial joints (Gray, 1973).
It has long been believed that once articular cartilage is damaged it is incapable of repair (Hunter, 1743; Paget, 1853). This characteristic is attributed to the lack of blood and nervous supply to this type of connective tissue (Barnett, 1961; Brower and Akahosi, 1962; McKibbin, 1973). In vascularized tissues, there are three well-documented phases of response to injury which are lacking in articular cartilage following injury (Paget, 1853; Mankin, 1982). Due to the vascular supply of the subchondral bone, lesions which fracture it undergo the three normal stages of repair: necrosis, inflammation, and healing. The repair tissue in these defects, however, is fibrocartilage containing type I collagen which does not have the same biomechanical/biochemical character as hyaline cartilage which is comprised of predominantly type II collagen (Campbell, 1969).
Cartilage defects which do not fracture the subchondral plate can progress to necrosis but not inflammation or repair. Studies of these defects have indicated that initially there is a heightened metabolic activity demonstrated by an increased uptake by chondrocytes of sulphate and glycine (Mankin and Lipiello, 1969; Mankin, 1982). However, these attempts at repair are short-lived and after one week the increased synthesis of matrix products usually returns to normal levels of activity. Long term observation shows that these lesions do not heal significantly (Mankin,1982).
The question of whether articular cartilage has an intrinsic ability to heal after injury has not been adequately answered (Hunter, 1743; Redfern, 1851; Paget, 1853; Mankin, 1982; Sokoloff, 1984). In reviewing the literature, it becomes apparent that the results reported by various investigators have been far from unequivocal. Two possible mechanisms may exist by which repair may be initiated (Sokoloff, 1974, 1978). First is replication of chondrocytes in the regions adjacent to a defect, also termed "intrinsic" repair. Second, is metaplasia of cartilage from other para-articular connective tissue within the joint capsule such as synovium and subchondral bone, termed "extrinsic" repair. To what degree either of these mechanisms can contribute to the total restoration of a joint surface is unknown.
It is possible to characterize three basic types of defects in articular cartilage after injury. A type I defect consists of scarification of the articular surface, similar to the fibrillar changes that can be seen in early degenerative joint disease (Meachim, 1963; Mankin, 1982). A type II defect is comparable to the erosive thinning that occurs in rheumatoid arthritis and in osteoarthritis (DePalma et al., 1963, 1966; Green, 1977, Mankin, 1982). A type II defect penetrates all layers of cartilage down to but not including the subchondral plate. A type III defect is a full-thickness cartilage defect with some loss of subchondral bone. Type III defects can be compared to osteochondral fractures or drill defects.
There is confusion in the literature over the definition of "full-thickness cartilage defects" both clinically and experimentally. Some authors define full-thickness defects as being those that include fracturing of the subchondral plate; others define it as penetration of all layers of cartilage without violation of the subchondral bony plate (Kennedy et al., 1967; Salter, 1980; Rubak et al., 1982A, 1982B; Johnson-Nurse and Dandy, 1985). Injuries producing types I and II defects exhibit a similar minimum degree of repair. Due to the vascular supply of the subchondral bone, type III lesions undergo the three normal stages of repair: necrosis, inflammation, and healing.
The idea of allografting to heal lesions in articular cartilage goes back over a century (Hunter, 1943; Bert, 1865; Zhan, 1877; Tizzoni, 1878; Dupertius, 1941). Early investigators also observed that any cartilaginous tissue produced following an injury was not hyaline cartilage but fibrocartilage (Redfern, 1851; Paget, 1953).
Two separate techniques are currently used in cartilage transplantation research: (1) the transplantation of osteochondral grafts, and (2) the transplantation of chondrocytes. Numerous attempts have been made to transplant whole or partial joints with mixed results (DePalma et al., 1963; Seligman, 1972). The availability of clostridial collagenase to enzymatically isolate chondrocytes from their matrix made it possible to attempt culture as well as transplantation of chondrocytes (Smith, 1965; Manning and Bonner, 1967). Subsequently, many investigators have attempted to successfully heal cartilage defects using the method of chondrocyte transplantation. Earlier studies reported using chondrocytes isolated from epiphyseal plates (Chesterman and Smith, 1968; Bentley and Greer, 1971; Green, 1977; Bentley et al., 1978) as well as articular chondrocytes (Chesterman and Smith, 1968; Green, 1977).
It was believed that the epiphyseal cells would perform better as grafts due to their heightened metabolism. However, these grafts generally did not heal well, probably due to their fate of eventual hypertrophy, necrosis, and calcification (Bloom and Fawcett, 1975). Techniques using drills to create defects (Bentley and Greer, 1971; Bentley et al., 1978) may suffer from two shortcomings: (1) Defects are not reproducibly made to the same depth. Violation of the subchondral plate may therefore have occurred. (2) Heat dissipated by drill friction may cause local necrosis in the adjacent matrix.
In the numerous reports of chondrocyte transplantation since the introduction of collagenase, none has addressed a method for internal surgical fixation of the grafted cells in vivo. This could possibly account for the reported low incidence of healing. It would seem improbable that a graft of free cells would remain within a defect in the joint environment without diffusing outward. Another problem remains in that freshly isolated cells, being relatively small in number and having recently been traumatized by the rigors of enzyme digestion, would have a minimum potential to reconstitute defects. In addition, some authors provide results distinguishing between autograft and allograft transplants (DePalma et al., 1966), while others do not (Chesterman and Smith, 1968; Green, 1971, 1977; Green and Ferguson, 1975).
Injury to articular cartilage is probably more frequent than is diagnosed either by clinical or radiographic examination. It has been noted that localized articular cartilage lesions were found in 30 to 60% of acute and chronic knee injuries in humans (Clancy et al., 1983) and occur in a variety of injuries such as blunt trauma, fractures, and dislocations. At present, the available treatment for these lesions, which includes excision and drilling, articular cartilage debridement, arthroscopic shaving, and abrasion arthroplasty, shows that optional resurfacing in human patients is not achieved because only fibrocartilaginous repair is the result (Magnuson, 1941; Henche, 1967; Insall, 1967; Mitchell and Shepard, 1978; Johnson, 1981).